title: Microfluidic on-chip capture-cycloaddition reaction to reversibly immobilize small molecules or multi-component structures for biosensor applications.
abstract: Methods for rapid surface immobilization of bioactive small molecules with control over orientation and immobilization density are highly desirable for biosensor and microarray applications. In this Study, we use a highly efficient covalent bioorthogonal [4+2] cycloaddition reaction between trans-cyclooctene (TCO) and 1,2,4,5-tetrazine (Tz) to enable the microfluidic immobilization of TCO/Tz-derivatized molecules. We monitor the process in real-time under continuous flow conditions using surface plasmon resonance (SPR). To enable reversible immobilization and extend the experimental range of the sensor surface, we combine a non-covalent antigen-antibody capture component with the cycloaddition reaction. By alternately presenting TCO or Tz moieties to the sensor surface, multiple capture-cycloaddition processes are now possible on one sensor surface for on-chip assembly and interaction studies of a variety of multi-component structures. We illustrate this method with two different immobilization experiments on a biosensor chip; a small molecule, AP1497 that binds FK506-binding protein 12 (FKBP12); and the same small molecule as part of an immobilized and in situ-functionalized nanoparticle.
authors.affiliation: Center for Systems Biology, Massachusetts General Hospital.
authors.forename: Carlos
authors.initials: C
authors.lastname: Tassa
authors.forename: Monty
authors.initials: M
authors.lastname: Liong
authors.forename: Scott
authors.initials: S
authors.lastname: Hilderbrand
authors.forename: Jason E
authors.initials: JE
authors.lastname: Sandler
authors.forename: Thomas
authors.initials: T
authors.lastname: Reiner
authors.forename: Edmund J
authors.initials: EJ
authors.lastname: Keliher
authors.forename: Ralph
authors.initials: R
authors.lastname: Weissleder
authors.forename: Stanley Y
authors.initials: SY
authors.lastname: Shaw
mesh_terms: D015374:Biosensing Techniques / Q000295:instrumentation / Q000379:methods*; D003500:Cyclization; D034242:Cyclooctanes / Q000737:chemistry*; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation / Q000379:methods*; D053758:Nanoparticles / Q000737:chemistry; D020349:Surface Plasmon Resonance; D013777:Tetrazoles / Q000737:chemistry*
chemical_list: D034242:Cyclooctanes; D013777:Tetrazoles